Parental Lines Research Articles

Novel HER2-targeted therapy to overcome trastuzumab resistance in HER2-amplified gastric cancer

Trastuzumab is used to treat HER2-amplified metastatic gastric cancer; however, most patients become trastuzumab-resistant within a year. Knowledge of the mechanisms underlying trastuzumab resistance is required to overcome this limitation. Here, we aimed to elucidate this resistance mechanism using four trastuzumab-resistant (TR) cell lines and investigate the efficacy of HER2-targeted therapies to overcome treatment resistance. Each TR cell line had different phenotypic characteristics. Interestingly, HER2 expression remained as high as the parental cell lines in TR cell lines, suggesting that HER2-targeted agents were still useful. As expected, three tyrosine kinase inhibitors (lapatinib, neratinib, and tucatinib) and one antibody–drug conjugate (trastuzumab deruxtecan: T-DXd) exhibited good antitumor effects against TR cell lines. We further investigated the potential biological mechanism of T-DXd. When treated with trastuzumab or T-DXd, HER2 or its downstream signals were disrupted in parental cell lines, but not in TR cell lines. Moreover, T-DXd induced the expression of pH2A.X and cPARP and caused cell cycle arrest in the S or G2-M phase in TR cell lines. T-DXd showed promising antitumor activity in both parental and TR cell lines, suggesting that it is a potential candidate for overcoming trastuzumab resistance.

Genetic study of fruit quality traits in watermelon (<i>Citrullus lanatus</i>) prebred line derived from <i>Citrullus lanatus</i> var. <i>citroid</i>

Fruit quality is most important for commercially cultivated watermelon [Citrullus lanatus (Thunb.) Matsum and Nakai] cultivars and introgression of resistance gene from wild or related species is a major challenge and imperative to study the inheritance of internal and external fruit quality traits for utilizing in resistance breeding programme. Present study was carried out at the research farm of ICAR-Indian Agricultural Research Institute, New Delhi during summer 2020 to 2022 involving 4 seasons for identifying source of resistance, developing F1 and other subsequent generations (F2, BC1P1 and BC1P2) and evaluation of parental lines along with breeding population in watermelon. The resistant line DWM-45 was used as a female parent and crossed with commercially popular sweet watermelon variety Arka Manik to produce F1 and other generation to study the inheritance of internal and external fruit quality traits. Based on the segregation pattern of progeny it is concluded that fruit shape, rind pattern, seed coat colour and colour of flesh at margin near rind and at carpellar region followed monogenic inheritance. Furthermore, our findings revealed a complex genetic control with epistatic interactions for other fruit quality traits such as fruit length, fruit width, fruit weight and total soluble solids. This study provides valuable insights into the genetic basis of various fruit quality traits in watermelon which can have practical applications in breeding programmes, facilitating the development of improved watermelon varieties with desirable fruit characteristics.

Impact of Submergence1 (Sub1) Locus in Genetic Background of Modern Rice Varieties (Oryza sativa L.)

Aim: To improve submergence stress tolerance of modern rice varieties (ADT36 and ADT37) through selection of superior rice lines.
 Study Design: Backcross method.
 Place and Duration of Study: This effort was taken in Nethouse of Kandaswami Kandar’s College, Namakkal District, Tamil Nadu, India. This process was completed in four years from 2019 to 2022.
 Methodology: Here, widely cultivated two short-term rice varieties (ADT36 and ADT37) were selected to improve submergence tolerance based on polymorphism at phenotypic and genotypic level. Then, they were improved through backcrossing with respective recurrent parent by selection of superior rice lines compared to parental lines under submergence condition. Likewise, this process was advanced up to BC3F2 generation.
 Results: In this study, a number of two NILs for ADT36 rice variety (ADT36- F12-5-19.18 and ADT36- F15-2-5.17) and five NILs for ADT37 rice variety (ADT37- F113-27-5.2, ADT37- F113-27-5.5, ADT37- F115-2-1.10, ADT37- F115-2-1.14 and ADT37- F115-2-2.14) are selected as superior lines.Selected rice lines exhibited more than 60% submergence tolerance than donor parent. This result indicates the adaptation of Sub1 locus into new genetic background of ADT36 and ADT37 rice variety under submergence condition.
 Conclusion: Improvement process of rice lines under stress condition results in identification of highly active rice lines. Thus, selected submergence tolerant rice lines would be a boon to the rice farmers who are facing crop lose due to frequently occurring cyclones in the Cauvery delta region.

Effect of provenance on flowering behaviour of parental lines of sunflower(Helianthus annuus L.) Hybrid-KBSH-78

The study revealed that sowing of parental lines of sunflower hybrid KBSH-78 at different locations had no significant influence on plant growth specially flowering behaviour and seed yield. However, 50% flowering was observed at 57 DAS in male parent and 62 DAS in female parent i.e., female parent is late in 50% flowering by 5 days compared to male parent. In order to achieve the better synchronization between parental lines and also to enhance seed yield of the seed parent, male parent has to be sown late with two staggered sowings of 3rd and 5th day after sowing of female parent, irrespective of the seed production locations.

Aeroplane wing, a new recessive autosomal phenotypic marker in the malaria vector, Anopheles stephensi Liston

A novel and distinct mutant with a phenotype, aeroplane wing (ae) is reported for the first time in the urban malaria vector Anopheles stephensi. The main aim of this study was to establish the mode of inheritance of the ae gene performing genetic crossings between the mutants and wild types. These mutants show extended open wings that are visible to naked eyes in both the sexes. Mutants were first noticed in a nutritionally stressed isofemale colony. Strategic genetic crosses revealed that the ae gene is a recessive, autosomal, and monogenic trait having full penetrance with uniform expression in its adult stage. Egg morphometric analysis confirmed that these mutants were intermediate variant. No significant differences were observed in the wing venation and size of ae mutants compared to their control parental lines. Further cytogenetic analysis on the ovarian polytene chromosome of ae mutant showed an inversion (3Li) on the 3L arm like its parental line. This ae mutant would be a prominent marker and could be useful to study the functions of related specific genes within its genome.

Variability analysis for agronomic traits in Indian mustard germplasm

Eighty four inbred lines of mustard were evaluated during 2021-22 at ICAR-Directorate of Rapeseed Mustard Research, Bharatpur. Analysis of data revealed significant variability for all observed traits except days to flower senescence and days to maturity. Highest variability was recorded for biological yield followed by plant height and Main shoot length (cm). Days to flowering senescence and days to maturity had low variability observed. Plant height and1000-seed weight was observed moderate variability. Promising lines were identified for number of days to flowering initiation (<46 days), long main shoot length (>95cm), number of seeds per siliqua (>18), 1000- seed weight (>6.5g).These promising lines shall further be used in hybrid breeding programme as parental lines, as well as in varietal development programme.

Evaluation of Castor parental lines for root growth (Poly Bags)

Twelve castor parental lines were grown in poly bags to see the root and shoot growth till 90 DAS during kharif, 2020 in CRBD with 4 replications and 3 plants per replication. Total dry matter (TDM) showed strong positive correlation (>0.70) with all shoot and root traits. Among the studied parental lines, IPC-41, IPC-43, DPC-22 recorded good crop growth (stem girth, root length, root volume), root, stem dry weight and total dry matter (TDM).

Generation and characterization of a human induced pluripotent stem cell line heterozygous for a NOTCH1 mutation (NCHi014-A)

NOTCH1 signaling is crucial for cardiovascular development. Numerous studies have identified heterozygous NOTCH1 loss of function and missense variants associated with a spectrum of congenital heart diseases (CHD). We generated induced pluripotent stem cells (iPSC) from a healthy individual to develop a model for NOTCH1+/- iPSC to study the molecular pathogenesis of CHD. NOTCH1+/-iPSC (NCHi014-A) have normal morphology and karyotype, are identical to the parental cell line, express pluripotency markers and have the ability to differentiate to the three germ layers. NOTCH1+/- iPSC can be used as a tool to study the cellular and molecular mechanisms underlying NOTCH1-associated human CHD.

Phospholipid scramblase 1 is involved in immunogenic cell death and contributes to dendritic cell–based vaccine efficiency to elicit antitumor immune response in vitro

Background aimsWhole tumor cell lysates (TCLs) obtained from cancer cells previously killed by treatments able to promote immunogenic cell death (ICD) can be efficiently used as a source of tumor-associated antigens for the development of highly efficient dendritic cell (DC)-based vaccines. Herein, the potential role of the interferon (IFN)-inducible protein phospholipid scramblase 1 (PLSCR1) in influencing immunogenic features of dying cancer cells and in enhancing DC-based vaccine efficiency was investigated. MethodsPLSCR1 expression was evaluated in different mantle-cell lymphoma (MCL) cell lines following ICD induction by 9-cis-retinoic acid (RA)/IFN-α combination, and commercial kinase inhibitor was used to identify the signaling pathway involved in its upregulation. A Mino cell line ectopically expressing PLSCR1 was generated to investigate the potential involvement of this protein in modulating ICD features. Whole TCLs obtained from Mino overexpressing PLSCR1 were used for DC loading, and loaded DCs were employed for generation of tumor antigen-specific cytotoxic T lymphocytes. ResultsThe ICD inducer RA/IFN-α combination promoted PLSCR1 expression through STAT1 activation. PLSCR1 upregulation favored pro-apoptotic effects of RA/IFN-α treatment and enhanced the exposure of calreticulin on cell surface. Moreover, DCs loaded with TCLs obtained from Mino ectopically expressing PLSCR1 elicited in vitro greater T-cell–mediated antitumor responses compared with DCs loaded with TCLs derived from Mino infected with empty vector or the parental cell line. Conversely, PLSCR1 knock-down inhibited the stimulating activity of DCs loaded with RA/IFN-α–treated TCLs to elicit cyclin D1 peptide-specific cytotoxic T lymphocytes. ConclusionsOur results indicate that PLSCR1 improved ICD-associated calreticulin exposure induced by RA/IFN-α and was clearly involved in DC-based vaccine efficiency as well, suggesting a potential contribution in the control of pathways associated to DC activation, possibly including those involved in antigen uptake and concomitant antitumor immune response activation.

Development of wilt resistant parental lines through mutagenesis in castor(Ricinus communis L.)

The present study was taken up to diversify the pistillate and monoecious lines to be used in hybridization programme. Mutagenesis was carried out using the chemical mutagen ethyl methane sulfonate (EMS) to induce mutation in popular pistillate line DPC 9 and land race Rasipuram local. The segregating mutant population was evaluated for their wilt resistance by screening in wilt sick plot. Plant to progeny row was continued up to M5 generation by selecting superior progenies in terms of yield and wilt resistance. Skewness and kurtosis indicated the presence of gene interactions for most of the traits studied. The pistillate progeny M5 29-11-5 (YRCP 2) and the monoecious progeny M4 106-2-3(YRCS 1904) were found to be the promising one and this could be further utilized as parents in hybrid development process.

Occurrence of castor diseases in Tamil Nadu and screening of castor parental lines for wilt resistance

Castor is an important industrial oilseed crop. The survey conducted during 2021-2022 in castor growing area of Tamil Nadu indicated that wilt and gray mold are the major disease. The wilt disease incidence ranged from 10.2 to 25.4 per cent and gray mold disease severity ranged from 37.8 to 62.2 PDI. The screening of castor parental line for wilt resistance in wilt epiphytic condition revealed BCS 3, ICS 253, ICS 309 and IPC 46 found promising for wilt resistance and they can be utilized for the development of wilt resistant castor hybrids.

Identification of resistant sources against Fusarium wilt of safflower

A field evaluation of 37 advanced generation breeding material and parental lines of safflower was carried out under wilt sick plot condition at Solapur, Tandur and IIOR, Hyderabad for identifying resistant sources against Fusarium wilt. Four genotypes viz., GMU-5032, GMU-4109, SAF-19-20, and SAF-19-09 were found resistant to Fusarium wilt at all three locations. These resistant sources can be used in breeding programme for development of a variety resistant to Fusarium wilt.

Enhancement of oil content in safflower (Carthamus tinctorious L.)

A systematic breeding approach is applied to increase the oil content in safflower lines by crossing between elite GMU lines and Mexican lines. The range of oil content in elite GMU parental lines were 33-35% were as in Mexican lines the average oil content was 39%. The crosses were made, through transgressive segregation the oil content in F5-F6 generations was achieved up to 42%.

SSR-marker assisted evaluation of genetic diversity in local and exotic pigeonpea cultivars in Benin for parental genotypes selection

Pigeonpea production is limited in Benin due to the undesirable agronomic and organoleptic characteristics of cultivated landraces. For a breeding program, some exotic cultivars were introduced for improving the available genepool in the country. The present study aims to assess the genetic diversity within a collection comprising of 32 local and 28 exotic cultivars, with the objective of selecting suitable parental lines for pigeonpea genetic improvement. All of the 60 accessions were genotyped with 14 SSR makers and a total of 38 alleles were detected with an average of 2.71 alleles per locus. The mean value of PIC (Polymorphism Information Content) and overall gene diversity were 0.37 and 0.43, respectively, while the mean value of heterozygosity was 0.11, indicating moderate genetic diversity within the 60 accessions with a deficit of heterozygous genotypes. As expected, the diversity was higher in exotic cultivars than in local landraces. Based on the genetic relationship among accessions, the entire collection formed 3 clusters, one of which comprised four specific exotic accessions. Population structure analysis using model-based and DAPC (Discriminant Analysis for Principal Components) methods subdivided the 60 accessions into two major groups reflecting their geographic origins with a minimal number (02) of admixtures. This indicates significant genetic divergence between local and exotic cultivars. Analysis of Molecular Variance (AMOVA) results with an overall Fst value of 0.13 confirmed the significant genetic divergence between the two collections. The findings of this study will significantly benefit for pigeonpea breeding programs in Benin. For practical applications, selected exotic cultivars, particularly those within the specific cluster can immediately serve as donor parents in crossbreeding schemes with the local landraces, for developing new varieties.

Microsatellite loci-based genetic relatedness holds key to parental selection for Sesame (Sesamum indicum L.) improvement

Microsatellite-based DNA-level diversity and genetic relatedness assessments empower parental selection in sesame crop improvement. A panel of 70 sesame varieties that were analyzed for 40 microsatellite loci got grouped into 5 distinct cluster based on Jaccard’s coefficient at genetic distance of 0.2. Nei coefficient analysis deciphered the presence of the maximum genetic distancebetween cluster 1 and cluster 5. The members of cluster 1 can be crossed with that of cluster 5 to get better allelic combinations. The findings of the present help selecting parental lines from a panel of 70 varieties for sesame crop improvement program.

Standardization of screening technique for root rot disease (Macrophomina phaseolina) of castor

Root rot caused by Macrophomina phaseolina is a major disease of castor in endemic areas and under water stress conditions. Sick pot method was found better artificial inoculation method for screening of castor genotypes than stem tape inoculation method and sorghum grown inoculum of M. phaseolina @ 35 g/kg soil standardized for sick pot method. In both inoculation methods, parental lines ICS-303, ICS-304, ICS-305, ICS-319, DPC-22, and IPC-46 recorded <10% root rot incidence, while ICS-298, ICS-312, and ICS-321 recorded <20% root rot incidence.

Inheritance and stacking effect of mutant ALS genes in Schoenoplectiella juncoides (Roxb.) Lye (Cyperaceae)

Schoenoplectiella juncoides, a noxious sedge weed in Japanese rice paddy, has two ALS genes, and ALS-inhibitor-resistant plants have a mutation in one of the ALS genes. The authors aimed (a) to quantitate the effect of the number of mutant alleles of ALS genes on whole-plant resistance of S. juncoides and (b) to clarify a mode of inheritance of the resistance by investigating resistance levels of the progenies of a hybrid between two S. juncoides plants with Trp574Leu substitution in different ALS. A dose-response analysis on the parental lines and the F1 population suggested that the two ALS genes contribute equally to whole-plant resistant levels. A dose-response study on the F2 population indicated that it could be classified into five groups based on the sensitivities to metsulfuron-methyl. The five groups (in ascending order of resistance levels) were considered to have zero, one, two, three, and four mutant alleles. The stacking effect of mutant alleles on resistance enhancement was more significant when the number of mutant alleles was low than when it was high; in other words, each additional mutant allele stacking increases plant resistance, but the effect saturates as the number of mutant alleles increases. A chi-square test supported that the segregation ratio of the five groups corresponds to 1:4:6:4:1 of Mendelian independence for the two ALS loci.

Inbred-hybrid relationship and heterosis on agronomic traits in bi-color sweet and waxy corn

Abstract. Patthawaro P, Dermail A, Chankaew S, Simla S, Lomthaisong K, Lertrat K, Suriharn K. 2023. Inbred-hybrid relationship and heterosis on agronomic traits in bi-color sweet and waxy corn. Biodiversitas 24: 6351-6359. Maize hybrids become more acceptable than other cultivar types since they promise high yield and uniformity by exploiting the heterosis advantages. Numerous lines are to be crossed to obtain the promising hybrids, and the progenies must be tested in multi-environment trials. Handling large number of genotypes in routine work is resource intensive; thus, the question arises if hybrid prediction based on the mid-parent (MP) value is feasible to reduce the workload. This study aimed to investigate the reliability of MP values in hybrid prediction through simple linear correlation and to estimate the magnitude of heterosis on given traits in sweet-waxy corn. Six parental lines and 10 F1 hybrids were evaluated in Khon Kaen in the dry and rainy seasons (2021/2022). Genotype and the interaction between genotype and season were significant for all observed traits. Waxy corn genotype 8A3-B was a good combiner for yield and yield components. Heterosis was trait-dependent, and it could imply parental adaptation when the estimation was done in contrasting environments. We found that the MP value can be used to predict the hybrid performance for flowering times only, but this approach was not effective for the rest agronomic traits. Since commonly, maize improvements are regarding multiple favorable traits, the strategy to use MP as single factor in hybrid prediction was not reliable. Other approaches, either SCA alone or the sum of mid-parent GCA and SCA, could be implemented for hybrid prediction in future studies.

Knockout of IRF3 and IRF7 genes by CRISPR/Cas9 technology enhances porcine virus replication in the swine testicular (ST) cell line.

Antiviral vaccines for pig diseases are essential to prevent epidemic outbreaks. However, their production is often hindered by inefficient manufacturing processes that yield lower quantities of the vaccine. To accelerate the progress of various areas of bioproduction, we have considered the necessity of enhancing viral replication efficiency by optimizing ST (swine testicular) cell lines that are commonly utilized in virus manufacturing. CRISPR/Cas9 gene-editing technology were utilized to create IRF3 or IRF7 knockout cell lines that facilitate high-titer viral stock production. Compared to the parental cell lines, the ST IRF3/7 KO cell line displayed a compromised antiviral response to a panel of viruses (Porcine epidemic diarrhea virus, Senecavirus A, Parainfluenza virus 5, and Getah virus), as evidenced by decreased expression of interferon and certain antiviral factors. The inhibition of these responses led to heightened viral replication and increased cytopathic effects, ultimately promoting apoptosis. As a result, the development of these cell lines offers a more efficient approach for biopharmaceutical companies to boost their virus production and reduce associated costs.

Nuclear isoform of RAPH1 interacts with FOXQ1 to promote aggressiveness and radioresistance in breast cancer

Radioresistance limits the efficacy of radiotherapy against breast cancer, especially the most lethal subtype of breast cancer, triple-negative breast cancer (TNBC). Epithelial-to-mesenchymal transition (EMT) is closely related to tumor radioresistance. In this work, we attempted to identify the key EMT-related transcription factor(s) that can induce radioresistance in breast cancer cells. A set of 44 EMT transcription factors were analyzed in parental and radioresistant TNBC cell lines. The function of FOXQ1, a differentially expressed transcription factor, was determined in TNBC radioresistance. FOXQ1-interacting proteins were identified by co-immunoprecipitation and mass spectrometry. Compared with parental cells, FOXQ1 was significantly upregulated in radioresistant TNBC cells. Silencing of FOXQ1 increased the radiosensitiviy of radioresistant TNBC cells both in vitro and in vivo. FOXQ1 associated with a nuclear isoform of RAPH1 (named RAPH1-i3) in radioresistant TNBC cells. Overexpression of RAPH1-i3 enhanced TNBC cell proliferation and migration, and most interestingly, induced radioresistance in parental TNBC cells when co-expressed with FOXQ1. Similar findings were observed in estrogen receptor-positive breast cancer cell lines that had co-expression of RAPH1-i3 and FOXQ1. Mechanistically, co-expression of RAPH1-i3 and FOXQ1 activated STAT3 signaling and increased the expression of CCND1, MCL1, Bcl-XL, and MMP2. Depletion of RAPH1-i3 impaired the radioresistance of radioresistant TNBC cells. Additionally, RAPH1-i3 upregulation was associated with advanced tumor stage and reduced disease-free survival in TNBC patients. These results collectively show that RAPH1-i3 interacts with FOXQ1 to promote breast cancer progression and radioresistance. RAPH1-i3 and FOXQ1 represent therapeutic targets for the treatment of breast cancer including TNBC.